Saethre-Chotzen syndrome is one of the most common autosomal dominant disorders of craniosynostosis in humans and is characterized by craniofacial and limb anomalies. The locus for Saethre-Chotzen syndrome maps to chromosome 7p21-p22. We have evaluated TWIST, a basic helix- loop-helix transcription factor, as a candidate gene for this condition because its expression pattern and mutant phenotypes in Drosophila and mouse are consistent with the Saethre-Chotzen phenotype. We mapped TWIST to human chromosome 7p21-p22, and mutational analysis on over 15 patients revealed nonsense, missense, insertion and deletion mutations (Howard et al. 1997). These mutations occur within the basic DNA binding, helix I and loop domains, or result in premature termination of the protein. In Apert syndrome, characterized by craniosynostosis and syndactyly of the hands and feet, recurrent mutations of the serine-proline dipeptide (either FGFR2 Ser252Trp or Pro253Arg) in the linker between the IgII and IgIII extracellular immunoglobulin-like domains, have been documented in more than 160 unrelated individuals. We performed prenatal diagnosis on a sporadic case with features consistent with this condition detected as early as the first trimester (Filkins et al. 1997). We have identified three novel FGFR2 mutations of this dipeptide, associated with distinct craniosynostotic phenotypes (Oldridge et al. 1997). A CG T mutation that predicts a Ser252Phe substitution, ascertained in a boy with mild Crouzon syndrome (craniosynostosis with normal limbs) is also present in three clinically normal members of his family. A CG->TT mutation that predicts a Ser252Phe substitution results in a phenotype consistent with Apert syndrome. Finally, a CGC->TCT mutation that predicts a double amino acid substitution (Ser252Phe and Pro253Ser) causes a Pfeiffer syndrome variant with mild craniosynostosis, broad thumbs and big toes, fixed extension of several digits, and only minimal cutaneous syndactyly. Treacher Collins syndrome is the most common of the human mandibulofacial dysostosis disorders. Recently, a partial TCOFl cDNA was identified and shown to contain mutations in TCS families. Here we present the entire exon/intron genomic structure and the complete coding sequence of TCOFl - TCOFl encodes a low complexity protein of 1411 amino acids, whose predicted protein structure reveals repeated motifs that mirror the organization of its exons. These motifs are shared with nucleolar trafficking proteins in other species and are predicted to be highly phosphorylated by casein kinase. Consistent with this, the full- length TCOFI protein sequence also contains putative nuclear and nucleolar localization signals. Throughout the open reading frame, we detected an additional eight mutations and several polymorphisms when over 50 TCS families were screened (Wise et al. 1997). We postulated that TCS results from defects in a nucleolar trafficking protein that is critically required during human craniofacial development.